H
err Gustav Druer, the Brno wine merchant has a problem and Brother Gregory has been asked to help.

You are to become his research assisitants and help him carry out a research investigation into the properties of microbes.

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Brother Gregory has been given a series of the microbes and been asked to determine the growth properties of each species to see where it can grow best.

He wants you, his research assistants, to try growing these microbe under a number of different environmental conditions and find out how fast they reproduce.

This investigation concerns the effect of temperature on the rate of growth, its upper and lower limits, and finding out at which temperature the microbes grow best.

Background

Microbes, even if they are supplied with all the necessary nutritional requirements, still may not grow.

Bacteria, single celled eukaryotes and other microbes, can only live and reproduce within a certain range of environmental conditions. Factors that can influence if or how microbes can grow are temperature, pH, dissolved gases, osmotic pressure and water availability.

Microbes, such as bacteria are more tolerant of environmental conditions than other organisms. However, each species has its own characteristic and particular range of values in which it grows and reproduces best.

Upper and Lower Values, and Temperature Range
Some species of microorganism can grow at temperatures as low as -10o C, and others at temperatures as high as 100o C - or higher. These upper and lower values are a function of cell metabolism. At lower temperatures molecules move slower, enzymes cannot mediate in chemical reactions, and eventually the viscosity of the cell interior brings all activity to a halt.

As the temperature increases, molecules move faster, enzymes speed up metabolism and cells rapidly increase in size. But, above a certain value all of these activities are proceeding at such high rates, enzymes start to denature, and the total effect is detrimental. Cellular growth ceases.

These boundary values define the maximum and minimum temperature at which life can exist (and grow). Each species of microbe has its own, unique upper and lower limit, which is a defining characteristic for that species.

Optimum Values

Somewhere between its characteristic upper and lower temperature limits, each species of microbe has a particular temperature at which it grows best. At this temperature all aspects of the cell metabolism function at their optimum values, the cell is able to rapidly increase in size and divide. When members of a species find themselves living at their optimum temperature, their growth rate is at its maximum value.

Psychrophiles

Bacteria that grow at temperatures in the range of -5oC to 30oC, with optimum temperatures between 10oC and 20oC, are called psychrophiles. These microbes have enzymes that catalyze best when the conditions are cold, and have cell membranes that remain fluid at these lower temperatures.

Sea water near the poles of the earth are rich in algae that can live below 0oC, and the psychrophilic bacteria that spoil milk, meat, vegetables and fruit are perfectly happy growing in a refrigerator. Although refrigeration is a good way of slowing down food spoilage, it cannot stop the growth of these bacteria.

Mesophiles

Microbes that grow at optimal temperatures in the range 20oC to 40oC, are called mesophilic. Important members of this group are those that live in and on warm blooded creatures, such as humans. Pathogenic bacteria and included here, as are symbiotic bacteria that live in the human body without harming it.

Thermophiles

Certain bacteria can live and grow at temperatures that exceed 50oC. These are thermophilic microbes that can tolerate the very harsh conditions decomposing organic material, the hot springs at Yellowstone National Park (where temperatures are at least 80oC to 85oC), or deep in the oceans by thermal vents bubbling up from the hot rocks just below the earth's crust.

Tools of the Trade

In these investigations, a tiny group of each microbe species are placed into a liquid, nutrient filled broth that has been sterilized (so no other bacteria will compete!). Usually this is in a special flask (called an "Erlenmeyer flask"), which is slowly shaken (to keep the microbes and nutrient at uniform distributions).

Each growing culture is carefully kept at the appropriate, and constant, temperature for the length of the experiment.

At regular intervals of time, small samples of the growing culture are taken from the flask and all reproduction of the microbes stopped by some poison or inhibitor (they can also be chilled or frozen). The size of the population at each time point is then determined.

The results of each of your investigations should be recorded as a table (a Table of Results). In these tables you should indicate the name of the microbe being studied, the temperature growth, and make an accurate record of either the growth data (growth curve), or the value of the generation time (generations per minute), as required.

The logatithmic value of the generations per hour should also be recorded on your table of results.

The results of each investigation should then be presented as a graph.

The horizontal axis of the graph should be the intervals of the different temperatures at which the microbes were grown. The vertical axis should represent the logarithmic value of the generations per hour determined for that sample.

This is called a Arrhenius plot.

The shape of these graphs or plots is characteristic for each species of microbe, but each organism will show an optimum temperature where growth proceeds most rapidly, and as the temperatures either exceed, or fall below that optimum, growth slows down. Above or below the maximum and minimum permissive temperatures, all growth stops.

Each investigation is carried out under a specific set of growth conditions.

A species of microbe is chosen first.

It is then necessary to chose a temperature. Use the thermometer sliding scale to set the chosen temperature. The value chosen will appear in the box.

For each temperature, click on "GROWTH" and record your results. In some investigations you will need to record the entire growth curve (data on extreme right), but for most investigations you only need to record the "generations per minute" and "log. value of the generations per hour".

Record all the temperatures and all the values where you see that the microbial species could grow at all. It is not necessary to record those values that occur when there is not microbial growth.